The hematopoietic system is one of the most complex tissues and includes a number of different cell types essential for survival. This process is highly regulated by the complex interplay of signaling pathways and changing sets of transcription factors that are present in hematopoietic cells. The discovery of a new class of transcribed ultraconserved genomic regions (UCRs) that are aberrantly expressed in human leukemias, prompt us to seek whether these UCRs are differentially expressed among the specific hematopoietic lineages and whether UCRs play a role in the differentiation of hematopoietic stem cells (HPCs) into specific lineages, in particular the erythroid and megakaryocytic lineages. Using a novel UCRs microarray platform we have characterized UCRs expression at different stages of hematopoietic differentiation and identified distinctive signatures associated with particular lineages. The overall goal of this proposal is to demonstrate that transcribed UCRs are functionally relevant during hematopoiesis. To achieve this goal we propose the following specific aims: 1) Specific Aim #1: To investigate the effects of ectopic over-expression or inhibition of UCRs expression on erythrocyte and megakaryocyte lineage differentiation "in vitro" and "in vivo" by using morphology, immunophenotype, colony assays and bone marrow transplant assay experiments and 2) Specific Aim #2: To investigate whether GATA-1 regulates UCRs expression by: 1) analyzing UCRs expression after GATA-1 activation using a conditional GATA-1 cell system;2) identifying UCRs transcription starting sites using Rapid Amplification of cDNA ends (RACE);3) identifying GATA-1 binding sites using Chip-sequencing and 4) performing promoter functional studies. This research has the potential to identify novel regulators of hematopoiesis and may give insights into basic biology of gene expression and cell fate determination. If successful, this research will uncover an unparalleled reservoir of molecular information on ultraconserved non-coding and coding RNAs in stem cell differentiation that will be extraordinarily valuable for the field. PUBLIC HEALTH RELEVANCE: This research has the potential to give insights into the basic mechanisms of blood formation and to identify novel regulators in this process. This new information will be extraordinarily valuable for the field. Potential benefits include: 1) direct insight into normal hematopoietic regulation will also be important to understand malfunction in leukemias and other hematologic disorders and 2) the pharmacological modulation of hematopoietic stem cells by targeting critical regulators could be used to treat a wide variety of blood cell disorders that arise from abnormalities in the red or platelet family.